1x10^8 cells were fixed in 1% formaldehyde (Sigma, 252549) in PBS for 15- 20 min at room temperature and quenched with 1/10th volume of 1.25 M glycine for 5 minutes at room temperature. Cells were collected at 1,000xg for 5 minutes, washed in PBS, pelleted at 1,000xg for 5 min and pellets were flash frozen in liquid nitrogen and stored at -80°C until use. Pellets were thawed on ice and resuspended in 10 ml lysis buffer 1 (50 mM HEPES, pH 7.5, 140 mM NaCl, 1 mM EDTA, 10% glycerol, 0.5% IGEPAL CA-630, 0.25% Triton X-100 with 5μl/ml Sigma 8340 protease inhibitor cocktail incubated on ice 10 minutes, pelleted 1,500xg and subsequently washed in lysis buffer 2 (10 mM Tris-HCl pH 8.0, 200 mM NaCl, 1 mM EDTA, 0.5 mM EGTA and 5ul/ml protease inhibitor) as with lysis buffer 1 before resuspending in 1 ml lysis buffer 3 (10 mM tris-HCl pH 8.0, 1 mM EDTA, 0.1% SDS and 5 ul/ml protease inhibitors) for sonication. Chromatin was sheared in a 1 ml milliTUBE with AFA fiber on a Covaris E220 using 10% duty factor for 2 min. The sheared chromatin concentration was estimated with Nanodrop at OD 260 and diluted to 1 mg/ml in ChIP Dilution Buffer (10% Triton X-100, 1M NaCl and 1% Sodium deoxycholate). 1 mg chromatin was combined with 4 μg hybridoma tissue culture supernatant and rotated overnight at 4°C. 40 μl protein A/G Dynabeads was added and incubated for 2-4 hr rotating at 4°C. Samples were washed 5 times with 1 ml RIPA buffer, 2 times with TE with 50 mM NaCl. Chromatin was eluted with 800 μl elution buffer (50 mM Tris pH 8.0, 1 mM EDTA, 0.1% SDS) for 30 min at 65°C shaking at 1,500 rpm in a ThermoMixer (Eppendorf). Supernatants were collected, digested with 20 μl of 20 mg/ml Proteinase K and incubated overnight at 65°C. DNA was purified with phenol chloroform extraction. 500 μl of the aqueous phase was precipitated with 20 μl 5M NaCl, 1.5 μg glycogen and 1 ml EtOH on ice for 1 hr or at -20°C overnight. Sequencing libraries were prepared with the KAPA HTP library prep kit (Roche) using 1-10 ng DNA and libraries were size selected with AMPure XP beads (Beckman Coulter).